This study explores the adoptive transfer of T cells targeting hepatitis B virus (HBV) antigens and the hydrodynamic delivery of plasmids to induce viral protein production in a mouse model. The methodology involves precise injection techniques to enhance plasmid uptake in liver cells, facilitating immune response activation against HBV.
Take T cells with receptors that bind to the antigens of hepatitis B virus, or HBV.
Inject the T cells into the tail vein of a mouse.
Allow the injected cells to spread in the liver.
To mimic virus infection, take plasmids carrying the HBV genome.
Inject the plasmids rapidly into the tail vein, directing the plasmids into the liver.
The force of the liquid permeabilizes the vessel endothelium and generates temporary pores in the cells.
The plasmids enter the cell and undergo RNA synthesis, producing mRNA that is used for viral protein production.
Some of these proteins act as viral antigens.
The antigen-presenting molecules carry the viral antigens to the cell surface, exposing them to the T cells.
These immune cells interact with the viral antigens and initiate mechanisms to eliminate the infected cells, inhibiting viral protein assembly and spreading.
For hydrodynamic HPV plasmid delivery through the tail vein, dilute 10 micrograms of HPV plasmid in an 8% equivalent of the body mass of PBS. Load the plasmid into one 3-milliliter syringe equipped with a 26-gauge half-inch needle per mouse.
Heat HHD mice under a heat lamp for five minutes to dilate the tail veins, and carefully pull a mouse into a plastic restrainer. Locate a dilated lateral tail vein in the middle third of the tail, and insert the needle parallel to the vein to deliver the entire volume of plasmid through the tail vein over a three to five-second period.
For viral antigen-specific iPSC-derived CD8-positive T cell adoptive cell transfer, collect the iPSC CD8-positive T cells on day 22 of culture as demonstrated and seed the cells onto a new 10-centimeter cell culture dish.
After 30 minutes in the cell culture incubator, filter the floating cells through a 70-micrometer nylon strainer, and count the viable cells. Dilute sells to a 1.5 x 107 cells per milliliter of PBS concentration.
Inject 200 microliters of cells into individual four to six-week-old HHD mice into the tail vein, as just demonstrated. On day 14 after cell transfer, perform a hydrodynamic HPV plasmid delivery as demonstrated.
繁體中文
